1. Field of the Invention
The invention relates to methods for the pretreatment of a lignin containing biomass to render the biomass amenable to digestion. Pretreatment comprises the addition of calcium hydroxide and water to the biomass to form a mixture, and maintaining the mixture at a relatively high temperature. Alternatively, an oxidizing agent, selected from the group consisting of oxygen and oxygen-containing gasses, may be added under pressure to the mixture. The invention also relates to the digested products of the pretreated biomass which includes useful feedstocks, fuels, and chemicals such as sugars, ketones, fatty acids and alcohols, and to a method for the recovery of calcium from the pretreated biomass.
2. Description of the Background
Biomass can be classified in three main categories: sugar-, starch- and cellulose-containing plants. Sugar-containing plants (e.g. sweet sorghum, sugarcane) and starch-containing plants (e.g. corn, rice, wheat, sweet potatoes) are primarily used as food sources. Cellulose-containing plants and waste products (e.g. grasses, wood, bagasse, straws) are the most abundant forms of biomass. Although they are not easily converted to useful products, a well engineered process to convert them to feedstock may potentially be economical since the costs of feedstock are much less than those of sugar- and starch-containing biomass.
Cellulose-containing materials are generally referred to as lignocellulosics because they contain cellulose (40% -60%), hemicellulose (20% -40%) and lignin (10% -25%). Non-woody biomass generally contains less than about 15-b 20% lignin. Cellulose, a glucose polymer, can be hydrolyzed to glucose using acid, enzymes or microbes. Glucose can serve as a feedstock for fuel alcohol and single-cell protein production. Microbial hydrolysis produces cellular biomass (single-cell protein) and metabolic waste products such as organic acids. Acid hydrolysis, although simple, produces many undesirable degradation products. Enzymatic hydrolysis is the cleanest and most preferred approach. However, production of enzymes, mainly cellulase and cellobiase, can be an expensive step. Apart from alcohol production, lignocellulose can be used as inexpensive cattle feed. Since raw lignocellulose cannot be easily digested by cattle, it must be processed to improve its digestibility before it can be fed to ruminants. Also, anaerobic fermentation using rumen microorganisms can produce low molecular weight volatile fatty acids.
Cellulose is the world's most abundant biological material. Approximately 40% to 45% of the dry weight of wood species is cellulose. The degree of polymerization ranges from 500 to 20,000. Cellulose molecules are completely linear, unbranched and have a strong tendency to form inter- and intra-molecular hydrogen bonds. Bundles of cellulose molecules are thus aggregated together to form microfibrils in which highly ordered (crystalline) regions alternate with less ordered (amorphous) regions. Microfibrils make fibrils and finally cellulose fibers. As a consequence of its fibrous structure and strong hydrogen bonds, cellulose has a very high tensile strength and is insoluble in most solvents.
Hemicellulose is the world's second most abundant carbohydrate and comprises about 20% to 30% of wood dry weight. Hemicelluloses, although originally believed to be intermediates in cellulose biosynthesis, are formed through biosynthetic routes different from cellulose. Hemicellulose are heteropolysaccharides and are formed by a variety of monomers. The most common monomers are glucose, galactose and mannose (the hexoses) and xylose and arabinose (the pentoses). Most hemicelluloses have a degree of polymerization of only 200. Hemicelluloses can be classified in three families, xylans, mannans and galactans, named for the backbone polymer.
Lignin is the world's most abundant non-carbohydrate biomaterial. It is a three dimensional macromolecule of enormously high molecular weight. Since its units are extensively cross-linked, it is difficult to define an individual molecule. Lignin provides strength by binding cellulose fibrils together. Being hydrophobic in nature, it prevents water loss from the vascular system and, being highly resistant to enzymatic degradation, it protects plants from insects and microbial attack.
Phenylpropane, an aromatic compound, is the basic structural unit of lignin. The monomers not only cross-link with each other, but also covalently bond to hemicellulose. A great constraint to cellulose and hemicellulose accessibility is the presence of lignin. It has been shown that decreased lignin content causes increased digestibility. Lignin can be removed by physical, chemical, or enzymatic treatments. It must be decomposed to smaller units that can be dissolved out of the cellulose matrix. There are several well developed pulping methods that disintegrate and remove lignin, leaving the cellulose fairly intact. Conventional pulping processes, such as Kraft and sulfite pulping, are too costly as bioconversion pretreatments. Also, economical use of the removed lignin is difficult because its chemical structure and size distribution are highly heterogeneous.
Another major deterrent to enzymatic cellulosic hydrolysis is the highly ordered molecular packing of its crystalline regions. Cellulolytic enzymes readily degrade the more accessible amorphous portions of cellulose, but are unable to attack the less accessible crystalline material. Thus, enzymatic hydrolysis rates increase with decreasing crystallinity index measured by X-ray diffraction methods.
The moisture content of cellulose fibers influences enzymatic degradation. Cellulosic materials are effectively protected from deterioration by enzymes or microbes provided the moisture content is maintained below a critical level characteristic of the material and the organism involved. In general, this critical level is slightly above the fiber saturation point, approximately 40% of dry weight. Moisture plays three major roles: (1) it swells the fibers by hydrating cellulose molecules, thus opening up the fine structure and increasing enzyme access, (2) it provides a diffusion medium for enzymes and for partial degradation products, and (3) it is added to cellulose during hydrolytic cleavage of the glycosidic links of each molecule.
The surface area of lignocellulose is another important factor that determines susceptibility to enzymatic degradation. It is important because contact between enzyme molecules and the cellulose surface is a prerequisite for hydrolysis to proceed. A few other factors that also influence susceptibility include size and diffusibility of enzyme molecules in relation to size and surface properties of capillaries, unit cell dimensions of cellulose molecules, and conformation and steric rigidity of hydro-glucose units.
To enhance susceptibility to enzymatic hydrolysis, lignocellulose pretreatment is an essential requirement. The heterogeneous enzymatic degradation of lignocellulosics is primarily governed by its structural features because (1) cellulose possesses a highly resistant crystalline structure, (2) the lignin surrounding the cellulose forms a physical barrier and (3) the sites available for enzymatic attack are limited. An ideal pretreatment, therefore, would reduce lignin content, with a concomitant reduction in crystallinity and increase in surface area. Pretreatment methods can be classified into physical, chemical, physicochemical, and biological, depending on the mode of action. The literature available on this subject is voluminous. The various pretreatment methods that have been used to increase cellulose digestibility are summarized in Table 1.
TABLE 1 ______________________________________ Methods Used for the Pretreatment of Lignocellulosics. Physical Chemical Physicochemical Biological ______________________________________ Ball-milling Alkali Steam explosion Fungi Two-roll milling Sodium hydroxide Ammonia Fiber Hammer milling Calcium hydroxide Explosion Colloid milling Ammonia High pressure Acid steaming Sulfuric acid High energy Hydrochloric acid radiation Hydrofluoric acid Pyrolysis Gas Chlorine dioxide Nitrogen dioxide Oxidizing agents Hydrogen peroxide Ozone Cellulose solvents Solvent extraction Ethanol-water extraction Benzene-ethanol extraction ______________________________________
Biological Pretreatments
Biological pretreatments employ fungi for microbial de-lignification to make cellulose more accessible. Major biological lignin degraders are the higher fungi, Ascomycetes and Basidiomycetes. Fungal degradation is a slow process and most fungi attack not only lignin, but cellulose also, thus resulting in a mixture of lignin fragments and sugars. Improvements may require developing more specific and efficient microbes.
Physical Pretreatments
Physical pretreatments can be classified in two general categories: mechanical (involving all types of milling) and nonmechanical (involving high-pressure steaming, high energy radiation and pyrolysis). During mechanical pretreatments, physical forces, (e.g. shearing, compressive forces) subdivide lignocellulose into finer particles. These physical forces reduce crystallinity, particle size and degree of polymerization and increase bulk density. These structural changes result in a material more susceptible to acid and enzymatic hydrolysis. However, due to enormously high operating costs associated with the high energy requirements, low yields and large time requirements, these mechanical pretreatments are not practical. Nonmechanical physical pretreatment methods also increase digestibility, but have similar disadvantages and thus are not economical for real processes.
Physicochemical Pretreatments
Steam explosion and Ammonia Fiber Explosion (AFEX) are the main physicochemical pretreatments. Steam explosion heats wetted lignocellulose to high temperatures (about 25.degree. C.) and releases the pressure instantly. Due to rapid decompression, which flashes the water trapped in fibers, physical size reduction occurs. The high temperatures remove acetic acid from hemicellulose, so this process results in some autohydrolysis of the biomass. These changes result in better digestibilities, but the severe conditions also produce degradation products that inhibit hydrolysis and fermentation. These products are removed by washing with water which results in a loss of water soluble hemicellulose. Thus, although digestibilities are improved, biomass degradation and protein denaturization limits the use of steam explosion.
The AFEX pretreatment process soaks lignocellulose in liquid ammonia at high pressure and then explosively releases the pressure. Pretreatment conditions (30.degree. C. -100.degree. C.) are less severe than steam explosion. An increase in accessible surface area coupled with reduced cellulose crystallinity (caused by ammonia contacting) result in increased enzymatic digestibility. However, use of ammonia (a hazardous chemical) and the high pressure release makes the process quite complex and energy intensive.
Chemical Pretreatments
Many chemical treatments have been used for lignin removal and destruction of the lignin crystalline structure. Of these chemicals, acids, gases, oxidizing agents, cellulose solvents, and solvent extraction agents, are all able to increase digestibility, but are not as popular as alkalis. Economics, simpler processes and less degradation favor alkalis as chemical pretreatment agents. However, most of these are process for paper pulping and involves the complete or nearly complete destruction of lignin, and a corresponding destruction of cellulose. Although unimportant in pulping, these pulping process are quite severe and not useful as pretreatments for biomass. Furthermore, the traditional pulping processes used by the paper industry are too expensive as lignocellulose pretreatment methods.
U.S. Pat. No. 4,644,060 to Chou is directed to the use of super-critical ammonia to increase lignocellulose digestibility.
U.S. Pat. Nos. 4,353,713 and 4,448,588 to Cheng are directed to the gasification of biomass or coal which is an endothermic process. These patents also relate to a method for adding the required thermal energy by reacting lime with carbon dioxide which is an exothermic reaction.
U.S. Pat. No. 4,391,671 to Azarniouch is directed to a method for calcining calcium carbonate in a rotary kiln. The reference relates to the paper/pulp industry where the calcium carbonate would be contaminated with waste biomass. The waste biomass is burned to provide the needed heat of reaction.
U.S. Pat. No. 4,356,196 to Hulquist is directed to treating biomass with ammonia.
U.S. Pat. No. 4,227,964 to Kerr is directed to the use of ammonia to promote the kinking of pulp fiber to increase paper strength, not to break down the fibers.
U.S. Pat. No. 4,087,317 to Roberts is directed to the use of lime and mechanical beating to convert pulp into a hydrated gel. There is no mention of lime recovery or enzymatically hydrolyzing the hydrated gel.
U.S. Pat. No. 4,064,276 to Conradsen directed to a process where biomass is covered with a tarp and then ammoniated with ammonia, which is allowed to dissipate into the atmosphere.
U.S. Pat. No. 3,939,286 to Jelks is directed to oxidizing biomass with high-pressure oxygen under elevated temperature and pressure in the presence of an acid catalyst, and a metal catalyst, ferric chloride, to break lignin bonds and to increase digestibility. The catalysts are described as essential to the process and calcium hydroxide is utilized as a neutralizing agent to adjust the resulting pH of the hydrolyzed biomass.
U.S. Pat. No. 3,878,304 to Moore is directed to production of slow-release nonprotein nitrogen in ruminant feeds. An amide, urea, is reacted with waste carbohydrates in the presence of an acid catalyst. The resulting material is pelleted and used as animal feed. Since the nitrogen is released slowly in the rumen, it is nontoxic to the animal.
U.S. Pat. No. 3,944,463 to Samuelson et al. is directed to a process for producing cellulose pulp of high brightness. The cellulose is pretreated with an alkaline compound at a temperature of between about 60.degree. C. to about 200.degree. C. so as to dissolve between 1 and 30% of the dry weight of the material in the pretreatment liquor. The pretreatment liquor preferably contains sodium carbonate, sodium bicarbonate or mixtures thereof, or possible sodium hydroxide.
U.S. Pat. No. 3,639,206 to Spruill is directed to the treatment of waste water effluent derived from a pulping process with calcium oxide or hydroxide to reduce the fiber and color content of the effluent.
U.S. Pat. No. 4,048,341 to Lagerstrom et al. is directed to a process for increasing the feed value of lignocellulosic material by contacting the material with an alkaline liquid, specifically, sodium hydroxide. The alkaline liquid, supplied in excess, is allowed to run off the material before any essential alkalization effect has been reached. After the liquid absorbed in the material has provided its effect, an acid solution is added to the material to neutralize the excess alkali. The reference does not disclose the interrelationship of temperature and time of alkali treatment, nor does it disclose the optimal amounts of the sodium hydroxide and water.
U.S. Pat. No. 4,182,780 to Lagerstrom et al. is directed to a process for increasing the feed value of lignocellulosic materials by alkali treatment and subsequent neutralization of the materials with an acid in a closed system under circulation of the treating agents.
U.S. Pat. No. 4,515,816 to Anthony is directed to a process in which lignocellulose is treated with dilute acid in an amount of about 1.5 to 2.5% of the dry weight of lignocellulose. The mixture is then stored at ambient conditions for 5 to 21 days in an air-free environment.
U.S. Pat. No. 4,842,877 to Tyson is directed to a process for the delignification of non-woody biomass (&lt;20% lignin). In this process, non-woody biomass is treated with a chelating agent, to prevent unnecessary oxidation, and maintained at high pH and high temperatures (150.degree. F. to 315.degree. F.) in the presence of hydrogen peroxide and pressurized oxygen. Hydrogen peroxide is stated to cause a reaction on the cell walls to allow the hemicellulose and lignin to solubilize and be removed through a subsequent hydrolysis process. Oxygen is added to initiate and accelerate the activation of hydrogen peroxide.
The conditions and results of studies reported in the literature using ammonia (gaseous, anhydrous liquid, or NH.sub.4 0H) and sodium hydroxide as pretreatment agents are listed in Table 2 and Table 3, respectively. The literature available on the use of these two chemicals to enhance lignocellulose digestibility of ruminant feeds, as well as for hydrolysis to glucose, is extensive. The literature on calcium hydroxide pretreatment processes is considerably less compared to that for sodium hydroxide and ammonia. The conditions and results of studies reported in the literature using calcium hydroxide are shown in Table 4.
TABLE 2 __________________________________________________________________________ Reported Ammoniation Conditions Type of Ammonia gNH.sub.3 /kg dry Effect on Reference Biomass State Temp. (.degree.C.) Time Pressure Particle Size biomass Digestibility __________________________________________________________________________ Villareal, Coastal Gaseous ambient -- atmosp. -- 40 Increase in 1988 Bermuda grass DIT, CP.sup.1 Waiss et al., Rice straws NH.sub.4 OH 160 1 h -- 0.64 cm 26/52 Increased.sup.2 1972 Waiss et al., Rice straws NH.sub.4 OH ambient 30 d atmosp. 0.64 cm 50 Increased.sup.1 1972 Millet et al., Aspen sawdust Liquid 30/60/90 1 h 155/360/725 -- -- Increased by 1970 psi 51%.sup.2 Millet et al., Aspen sawdust Gaseous 30 1/2 to 74 h 155 psi -- -- Increased by 1970 47%.sup.2 Brown et al., Limpo grass Gaseous ambient 30 d atmosp. 2.5 cm 20/30/40 Increased.sup.1 1987 Kellens et Wheat straw NH.sub.4 OH 29 21 d atmosp. 2 mm 50 Increased al., 1983 from 13% to 33%.sup.2 Kellens et Wheat straw Gaseous 6 44 d atmosp. 2.5 cm 50 Increased.sup.1 al., 1983 Hultquist, Alfalfa Liq./Gas. 20-30 30 m 70-165 psig 1/16-1/2 in. 5/20 Increased by 1982 50%.sup.2 Millet et al., Aspen sawdust Gaseous -- 2 h 70 psi -- -- Increased by 1975 46%.sup.1 Morris et al., Corn stover -- ambient -- atmosp. 1.3 cm 30 Increase in 1980 DIT, DE.sup.1 __________________________________________________________________________ .sup.1 In vivo. .sup.2 In vitro
TABLE 3 __________________________________________________________________________ Reported NaOH Treatment Conditions g NaOH/100 g NaOH/100 Effect on Reference Type of Biomass Temp. (.degree.C.) Time Particle Size g solution g biomass Digestibility __________________________________________________________________________ Moore et al., Cotton linters 30 1 h 40 mesh 1 20 No effect.sup.2 1972 Moore et al., Aspen 30 1 h 40 mesh 1 20 Increased from 1972 10% to 50%.sup.2 Millet et al., Different wood 25 1 h -- 1 20 Increased.sup.2 1970 samples Fient et al., Different wood 25 1 h, 2 h 40 mesh 0.5, 1 2-20 Increased.sup.2 1970 samples Baker et al, Aspen sawdust ambient 2 h 0.16 cm 0.5 5 Increased from 197570 41% to 52%.sup.1 Anderson et Ryegrass straw ambient 24 h 2.54 cm 0.5-8 7.5-120 Increased from al., 1973 33% to 90%.sup.2 Mandels et al., Bagasse 72 1 h 1/8" mesh 2 -- Increased.sup.2 1974 Mandels et al., Newspaper 70 90 m 1/8" mesh 2 100 Increased.sup.2 1974 Turner et al., Different grass -- 48 m 4 mm 3 -- Increased.sup.2 1990 samples __________________________________________________________________________ Notation used; .sup.1 In vivo, .sup.2 In vitro
TABLE 4 __________________________________________________________________________ Reported Ca(OH).sub.2 Treatment Conditions g water/g g Ca(OH).sub.2 /100 Effect on Reference Type of Biomass Temp. (.degree.C.) Time Particle Size solution g biomass Digestibility __________________________________________________________________________ Playne, 1984 Bagasse 20 8 d 2.25 0.87 12 to 30 Increased from 1972 19% to 72%.sup.2 Waller et al., Corn cobs ambient 14 d Ground 0.6 4 Improved 1975 digestibility.sup.1 Rounds et al., Corn cobs -- -- -- -- 4 No effect.sup.2 1974 Gharib et al., Poplar bark ambient 1 or 150 d 9.5 mm 0.6 4 to 16 Increased from 1975 30% to 52%.sup.2 Felix et al., Soyabean straw ambient/frozen 30 d Chopped 0.65 2 to 5 No effect.sup.1 1990 __________________________________________________________________________ Notation used; .sup.1 In vivo, .sup.2 In vitro
The references cited in the Tables and below are:
Anderson, D. C.; Ralston, A. T. J. Anim. Sci. 1973; 37, 148. PA0 Baker, A. J.; Millett, M. A.; Satter, L. D. ACS Symposium Series 1975; 10, 75. PA0 Brown, W. F.; Phillips, J. D.; Jones, D. B. J. Anim. Sci. 1987; 64, 1205. PA0 Dawish, A.; Galal, A. G. In Proc. Conf Anim. Feeds Trop. Subtrop. Origin 1975. PA0 Felix, A.; Hill, R. A.; Diarra, B. Anim. Prod. 1990; 51, 47. PA0 Feist, W. C.; Baker, A. J.; Tarkow, H. J. Anim. Sci. 1970; 30, 832. PA0 Gharib, F. H.; Meiske, J. C.; Goodrich, R. D.; El Serafy, A. M. J. Anim. Sci. 1975; 40(4), 734. PA0 Hulquist, J. H. U.S. Pat. No. 4,356,296; 1982. PA0 Kellens, R. D.; Herrera-Saldana, R.; Church, D. C. J. Anim. Sci. 1983; 56(4), 938. PA0 Mandels, M.; Hontz, J. R.; Kystrom, J. Biotech. Bioeng. 1974; 16, 1471. PA0 Millet, M. A. et al., J. Anim. Sci. 1970; 31(4), 781. PA0 Millet, M. A.; Baker, A. J.; Satter, L. D. In Biotech. Bioeng. Symp. 1975; 5, 193. PA0 Moore, W. E.; Effland, M. J.; Medeiros, J. E. J. Agr. Food Chem. 1972; 20(6), 1173. PA0 Morris, P. J.; Movat, D. N. Can. J. Anim. Sci. 1980; 60, 327. PA0 Playne, M. J. Biotech. Bioeng., 1984; 26, 426. PA0 Rounds, W.; Klopfenstein, T. J. Anim. Sci. 1974; 39, 251 (abst.). PA0 Turner, N. D.; Schelling, G. T.; Greene, L. W.; Byers, P. M. J. Prod. Agric. 1990; 3(1), 83. PA0 Villareal, E. R. Ph.D. Thesis Texas A&M Univ. College Station, TX 1988. PA0 Waiss, A. C. et al., J. Anim. Sci. 1972; 35(1). 109. PA0 Waller, J. C.; Klopfenstein, T. J Anim. Sci. 1975, 41 424 (abstract).
Playne (1984) investigated the effects of alkali treatment and steam explosion on baggase digestibility. The digestibility of untreated bagasse was 190 g organic matter (0M)/kg bagasse dry matter. It was raised to: 733 g organic matter by using NaOH (and also by using Ca(OH).sub.2 with Na.sub.2 CO.sub.3); to 430 g OM using NH.sub.3 ; and to 724 g OM using Ca(OH).sub.2. When Ca(OH).sub.2 alone was used, a high loading (about 180-300 g Ca(OH).sub.2 kg bagasse) was used. Gharib et al. (1975) used calcium oxide for in vitro evaluation of chemically treated poplar bark. They reported that calcium oxide increased in vitro true digestibility from 38% to 52% for a 150-day treatment, although little improvement was found for a 1-day treatment. Rounds and Klopfenstein (1974) studied the effects of NaOH, KOH, NH.sub.4 0H and Ca(OH).sub.2 on in vivo digestibility of corn cobs by feeding to lambs and on in vitro digestibility using an artificial rumen. Ca(OH).sub.2 alone was unable to increase the in vitro digestibility, although rations treated with Ca(OH).sub.2 +NaOH resulted in higher daily gain and feed efficiency for lambs. Waller and Riopfenstein (1975) used various combinations of NaOH, Ca(OH).sub.2 and NH.sub.4 0H for treating feed for lambs and heifers and reported that the highest daily gain and lowest feed/gain was obtained for the 3% NaOH +1% Ca(OH).sub.2 rations. Darwish and Galal (1975) used maize cobs treated with 1.5% Ca(OH).sub.2 in a milk production ration and found no significant change in milk output. Felix et al. (1990) evaluated the effects of ensiling and treating soya-bean straw with NAOH, Ca(OH).sub.2 and NH.sub.4 0H on ruminant digestibility. Results indicate that there was no significant improvement due to alkali treatment of dry and unensiled straw, although alkali treatment improved digestibility of ensiled straw.
Although the use of calcium hydroxide as a pretreatment agent has been demonstrated, considerably less work has been done employing this chemical compared to other alkalis. Most of the previous work has been performed by animal scientists trying to develop a very simple process to increase the lignocellulose digestibility of animal feed. All these studies were done at room temperature or below, at lower water loadings, for very long periods and without any mixing. These processes required very long treatment times which is very expensive since the reactors must be very large. There is thus a need to improve the currently existing methods of pretreating lignocellulose containing material to render it amenable to enzymatic digestion.